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System and method for rapid detection and characterization of bacterial colonies using forward light scattering

a technology of forward light scattering and bacterial colonies, applied in the field of bacteria colonies, can solve the problems of requiring a skilled technician, putting the public at risk, and using sophisticated and expensive molecular tools

Active Publication Date: 2006-08-03
PURDUE RES FOUND INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

[0009] By way of introduction only, in one embodiment a system and method of rapid detection and characterization of bacteria growing in colonies using forward light scattering is described. A bacterial colony may be illuminated with a laser beam, and the light scattered in the forward direction by the colony may be detected and analyzed. Automated analysis of the scattering light patterns provides rapid detection and characterization of a variety of strains of Listeria, Escherichia coli, and other bacteria and microorganisms.

Problems solved by technology

Pathogenic bacterial contamination in products not only put the public at risk, but also is costly to companies because of routine product testing, product recalls and liabilities.
However, the ability of these methods to differentiate live from dead bacteria and specificity of different strains of closely related species, such as Listeria, are of concern.
The assays for identification of the bacterial colonies require multiple steps, and / or use sophisticated and expensive molecular tools requiring a skilled technician to operate the tests.

Method used

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  • System and method for rapid detection and characterization of bacterial colonies using forward light scattering
  • System and method for rapid detection and characterization of bacterial colonies using forward light scattering
  • System and method for rapid detection and characterization of bacterial colonies using forward light scattering

Examples

Experimental program
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example 1

Colony Culture Conditions

[0035] The bacterial cultures listed in the tables were obtained from a frozen glycerol stock (−80° C.) from a collection at Purdue University. The bacteria were grown in brain heart infusion broth (BHI) (Difco, available from Becton Dickinson and Company, Franklin Lakes, N.J.) for 18-24 h at 37° C. The cultures were propagated twice in BHI overnight at 37° C. at 140 rpm in a shaker incubator (New Brunswick Scientific, Edison, N.J.). For individual colony growth, BHI broth supplemented with 1.5% agar (Difco) in Petri dishes was used.

[0036] Bacterial plates for light scattering experiments were prepared from cultures grown in BHI. The cultures were serially diluted in sterile 20 mM phosphate buffered saline, pH 7.4 (PBS) so that the dilutions when plated would produce about 30-50 colonies per plate. The diluents were evenly distributed on the surface of BHI agar plates, in duplicate, using a bent glass spreader and were incubated at 37° C. for 18-36 h in a ...

example 2

Comparison of Listeria Colony Characteristics

[0037] Several strains of Listeria were cultured according to the protocol described above and were placed in the system 100 where each colony was subjected to a laser diode having a wavelength of 635 nm and the forward light scatter was detected on a detector. The forward light scatter pattern was collected and analyzed according to the attributes shown in Table I above. The light scattering patterns of L. innocua, (B) L. monocytogenes, (C) L. ivanovii, (D) L. Welshimeri, (E) L. seeligeri, (F) L. grayi are shown in FIG. 4A-F.

[0038] Results of the analysis showed the following: The scattering pattern of L. innocua showed a clear central spot, isotropic background, and clear rings. The ring is relatively thick and diffuse, and the average intensity of the whole pattern appears to be high. The scattering pattern of L. monocytogenes indicated the presence of a diffused central spot with rings. The innermost ring is small and generally has ...

example 3

Mueller Matrix Characterization

[0039] As discussed above for the embodiment shown in FIG. 2, the full 16 elements of Mueller matrix measurement are possible to determine. The Mueller matrix shown below will be used to determine additional characteristics of the stains of bacteria analyzed in the system 200. The Mueller matrix is a 4×4 matrix, which relates incident stokes parameter Si0-Si3 and scattered stokes parameter Ss0-Ss3. (S ⁢S⁢ ⁢0S ⁢S⁢ ⁢1S ⁢S⁢ ⁢2S ⁢S⁢ ⁢3)=(M11M12M13M14M21M22M23M24M31M32M33M34M41M42M43M44)⁢(S ⁢S⁢ ⁢10S ⁢S⁢ ⁢11S ⁢S⁢ ⁢12S ⁢S⁢ ⁢13)

[0040] The matrix will be used to describe the polarization properties of scattered field with given incident field. Conversely, if the input and scattered stokes parameters are measured, the characteristic information of the objects may be extracted. Performing this measurement in two dimensional area, polarization characteristics of the bacterial colony will be obtained. In the embodiment shown in FIG. 1, the single intensity measur...

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Abstract

A system and a method of detecting and characterizing a bacterial colony are presented in which the results are determined within about 48 hours. The bacterial colony is disposed on a substrate and placed between a laser and detector. Light from the laser impinges upon and is scattered by the bacterial colony. The forward scattered light is detected by an optical detector. The signal from the optical detector is analyzed by an analyzer and displayed or supplied to a storage medium for review. As different strains of bacteria possess unique forward scattering fingerprints, the particular strain may be identified.

Description

[0001] This application claims the benefit of U.S. Provisional Application No. 60 / 631,666, filed Nov. 30, 2004.FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT [0002] This invention was made with Government support under Contract 1935-42000-035 awarded by the United States Department of Agriculture (USDA). The Government may have certain rights in the invention.BACKGROUND [0003] 1. Technical Field [0004] This invention relates to bacterial colonies, and more particularly, to a system and method of detecting and characterizing bacterial colonies. [0005] 2. Background Information [0006] Pathogenic bacterial contamination in products not only put the public at risk, but also is costly to companies because of routine product testing, product recalls and liabilities. For example, Listeria monocytogenes is one of the major foodborne pathogens with the highest mortality rate (20-28%) among the food pathogens. Traditional methods for detection and identification of bacterial contaminants, such a...

Claims

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Application Information

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Patent Type & Authority Applications(United States)
IPC IPC(8): C12Q1/04G02B21/00
CPCG01N21/21G01N21/47G01N21/4788G01N2021/4707
Inventor HIRLEMAN, EDWIN D. JR.GUO, SONGLINGBHUNIA, ARUN K.BAE, EUIWON
Owner PURDUE RES FOUND INC
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